1. Field of the Invention
This invention relates to compositions of matter that are reagents for preparing radiopharmaceuticals, methods for preparing radiopharmaceuticals using said reagents, the radiopharmaceuticals thus prepared, and methods for using such radiopharmaceuticals. In particular, the invention pertains to reagents that are monoamine, diamide, thiol-containing (MADAT) metal chelators, as well as conjugates between said metal chelating groups and a variety of specific targeting moieties. Also provided in one aspect of the invention are radiodiagnostic agents comprised of the metal chelators conjugated with specific targeting moieties and radiolabeled with gamma radiation-emitting radioisotopes. In another aspect are provided radiotherapeutic agents comprised of the metal chelators conjugated with specific targeting moieties and radiolabeled with cytotoxic radioisotopes. Kits comprising the radiopharmaceuticals of the invention and adjuvant agents for the preparation of the radiodiagnostic and radiotherapeutic agents of the invention are provided. Radiodiagnostic and radiotherapeutic methods for using the agents of the invention are also provided.
2. Description of the Prior Art
It is frequently clinically advantageous for a physician to be able to localize the site of a pathological condition in a patient using non-invasive means. Such pathological conditions include diseases of the lungs, heart, liver, kidneys, bones and brain, as well as cancer, thrombosis, pulmonary embolism, infection, inflammation and atherosclerosis.
In the field of nuclear medicine, certain pathological conditions are localized, or their extent is assessed, by detecting the distribution of small quantities of internally-administered radioactively labeled tracer compounds (called radiotracers or radiopharmaceuticals). Methods for detecting these radiopharmaceuticals are known generally as imaging or radioimaging methods.
In radioimaging, the radiolabel is a gamma-radiation emitting radionuclide and the radiotracer is located using a gamma-radiation detecting camera (this process is often referred to as gamma scintigraphy). The imaged site is detectable because the radiotracer is chosen either to localize at a pathological site or, alternatively, the radiotracer is chosen specifically not to localize at such pathological sites. In many situations it is a particular advantage to use a radiolabeled specific binding compound as a radiopharmaceutical, which localizes specifically to the pathological site in vivo.
A variety of radionuclides are known to be useful for radioimaging, including 67Ga, 99mTc (Tc-99m), 111In, 123I, 125I, and 169Yb. However, a number of factors must be considered for optimal radioimaging in humans. To maximize the efficiency of detection, a radionuclide that emits gamma energy in the 100 to 200 keV range is preferred. To minimize the absorbed radiation dose to the patient, the radioisotope should emit no alpha or beta particle radiation, and the physical half-life of the radionuclide should be as short as the imaging procedure will allow. To allow for examinations to be performed on any day and at any time of the day, it is advantageous to have a source of the radionuclide always available at the clinical site.
Tc-99m is the preferred radionuclide for scintigraphic imaging because it has no significant particulate radiation emissions and emits gamma radiation at about 140 keV, it has a physical half-life of 6 hours, and it is readily available on-site using a molybdenum-99/technetium-99m generator.
Other radionuclides used in the prior art are less advantageous than Tc-99m. This can be because the physical half-life of such radionuclides are longer, resulting in a greater amount of absorbed radiation dose to the patient (e.g., indium-111). Alternatively, the gamma radiation energies of such alternate radionuclides are significantly lower (e.g., iodine-125) or higher (e.g., iodine-131) than Tc-99m and are thereby inappropriate for quality scintigraphic imaging. Lastly, many disadvantageous radionuclides cannot be produced using an on-site generator.
Tc-99m is a transition metal that is advantageously chelated by a metal chelator or metal chelating moiety. Chelating moieties capable of binding Tc-99m can be covalently linked to various tarfeting molecules to provide a means for radiolabeling such targeting molecules. This is because the most commonly available chemical species of Tc-99m, pertechnetate (TcO4−), cannot bind directly to most targeting molecules strongly enough to be useful as a radiopharmaceutical. Complexing of Tc-99m with such radiolabel chelating moieties typically entails chemical reduction of the pertechnetate using a reducing agent such as stannous chloride.
The use of chelating agents for complexing Tc-99m is known in the prior art.
Byrne et al., U.S. Pat. No. 4,434,151 describe N2S2, homocysteine-containing chelating agents for Tc-99m.
Fritzberg, U.S. Pat. No. 4,444,690 describes a series of bisamide, bisthiol chelating agents for Tc-99m.
Byrne et al., U.S. Pat. No. 4,575,556 describe N2S2, homocysteine-containing chelating agents for Tc-99m.
Nosco et al., U.S. Pat. No. 4,925,650 describe Tc-99m chelating complexes.
Kondo et al., European Patent Application, Publication No. 483704 A1 disclose a process for preparing a Tc-99m complex with a mercapto-Gly-Gly-Gly moiety.
European Patent Application No. 84109831.2 describes bisamido, bisthiol Tc-99m ligands and salts thereof as renal function monitoring agents.
Burns et al., 1985, European Patent Application No. 85104959.3 describe bisamino, bisthiol compounds for preparing Tc-99m labeled brain imaging agents.
European Patent Application No. 86100360.6 describes dithiol, diamino, or diaminocarboxylic acids or amine complexes for making Tc-99m labeled imaging agents.
Kung et al., 1986, European Patent Application No. 86105920.2 describe bisamino, bisthiol compounds for making small, neutral Tc-99m brain imaging agents.
Bergstein et al., 1988, European Patent Application No. 88102252.9 describe bisamino, bisthiol compounds for making small, neutral Tc-99m imaging agents.
PCT International Patent Application Publication No. WO89/12625 describe bifunctional chelating complexes of bisamido, bisthiol ligands and salts thereof, for use as renal function monitoring agents.
Davison et al., 1981, Inorg. Chem. 20: 1629–1632 disclose oxotechnetium chelate complexes.
Fritzberg et al., 1982, J. Nucl. Med. 23: 592–598 disclose a Tc-99m chelating agent based on N,N′-bis(mercaptoacetyl)-2,3-diaminopropanoate.
Byrne et al., 1983, J. Nucl. Med. 24: P126 describe homocysteine-containing Tc-99m chelating agents.
Bryson et al., 1988, Inorg. Chem. 27: 2154–2161 describe neutral complexes of technetium-99 which are unstable to excess ligand.
Misra et al., 1989, Tet. Lett. 30: 1885–1888 describe bisamine bisthiol compounds for radiolabeling purposes.
Bryson et al., 1990, Inorg. Chem. 29: 2948–2951 describe chelators containing two amide groups, a thiol group and a substituted pyridine group, said chelators forming neutral Tc-99m complexes.
Taylor et al., 1990, J. Nucl. Med. 31: 885 (Abst.) describe a neutral Tc-99m complex for brain imaging.
Targeting molecules labeled with radioisotopes have been used as radiopharmaceuticals for both diagnostic and therapeutic purposes. A number of methods have been developed to label targeting molecules with radioisotopes. Particularly important are the isotopes of technetium for scintigraphic imaging and rhenium and tin for therapeutic purposes. Toward this end there have been many examples of chelating groups developed for labeling targeting molecules.
Hnatowich, U.S. Pat. No. 4,668,503 describe Tc-99m protein radiolabeling.
Tolman, U.S. Pat. No. 4,732,684 describe conjugation of targeting molecules and fragments of the metal-binding protein, metallothionein.
Ege et al., U.S. Pat. No. 4,832,940 teach radiolabeled peptides for imaging localized T-lymphocytes.
Nicolotti et al., U.S. Pat. No. 4,861,869 describe bifunctional coupling agents useful in forming conjugates with biological molecules such as antibodies.
Fritzberg et al., U.S. Pat. No. 4,965,392 describe various S-protected mercaptoacetylglycylglycine-based chelators for labeling proteins.
Morgan et al., U.S. Pat. No. 4,986,979 disclose methods for imaging sites for inflammation.
Fritzberg et al., U.S. Pat. No. 5,091,514 describe various S-protected mercaptoacetylglycylglycine-based chelators for labeling proteins.
Gustavson et al., U.S. Pat. No. 5,112,953 disclose Tc-99m chelating agents for radiolabeling proteins.
Kasina et al., U.S. Pat. No. 5,175,257 describe various combinations of targeting molecules and Tc-99m chelating groups.
Dean et al., U.S. Pat. No. 5,180,816 disclose methods for radiolabeling a protein with Tc-99m via a bifunctional chelating agent.
Flanagan et al., U.S. Pat. No. 5,248,764 describe conjugates between a radiolabel chelating moiety and atrial natiuretic factor-derived peptides.
Reno and Bottino, European Patent Application 87300426.1 disclose radiolabeling antibodies with Tc-99m.
Ranby et al., 1988, International Patent Application No. PCT/US88/02276 disclose a method for detecting fibrin deposits in an animal comprising covalently binding a radiolabeled compound to fibrin.
Dean et al., International Patent Application, Publication No. WO89/12625 teach bifunctional coupling agents for Tc-99m labeling of proteins.
Schoemaker et al., International Patent Application, Publication No. WO90/06323 disclose chimeric proteins comprising a metal-binding region.
Morgan et al., International Patent Application, Publication No. WO90/10463 disclose methods for imaging sites of inflammation.
Flanagan et al., European Patent Application No. 90306428.5 disclose Tc-99m labeling of synthetic peptide fragments via a set of organic chelating molecules.
Gustavson et al., International Patent Application, Publication No. WO91/09876 disclose Tc-99m chelating agents for radiolabeling proteins.
Rodwell et al., 1991, International Patent Application No. PCT/US91/03116 disclose conjugates of “molecular recognition units” with “effector domains.
Cox, International Patent Application No. PCT/US92/04559 discloses radiolabeled somatostatin derivatives containing two cysteine residues.
Rhodes et al., International Patent Application, Publication No. WO93/12819 teach peptides comprising metal ion-binding domains.
Lyle et al., International Patent Application, Publication No. WO93/15770 disclose Tc-99m chelators and peptides labeled with Tc-99m.
Coughlin et al, International Patent Application, Publication No. WO93/21151 disclose bifunctional chelating agents comprising thiourea groups for radiolabeling targeting molecules.
Knight et al., 1990, 37th Annual Meeting of the Society of Nuclear Medicine, Abstract #209, disclose thrombus imaging using Tc-99m labeled peptides.
Babich et al., 1993, J. Nucl. Med. 34: 1964–1974 describe Tc-99m labeled peptides comprising hydrazinonicotinamide derivatives.
Well-studied members of the class of chelating groups used for radiolabeling targeting molecules include diamide dithiols (DADS), also known as N2S2 chelators, and mercaptoacetyltriglycines (MAG3), also known as N3S chelators. Both of these types of chelating groups form stable chelators with technetium, and methods have been developed to link these chelators to targeting molecules.
Fritzberg, European Patent Application No. 853042255.4 disclose N/S complexes of technetium.
Fritzberg et al., European Patent Application No. 88104755.9 disclose N/S chelating agents.
In general, these methods require that the chelate be heated briefly (15 min) at 100° C. in solution to produce the stable chelate (see, for example, Fritzberg et al., 1986, European Patent Application No. 853042255.4). Since many targeting molecules such as peptides and carbohydrates are labile to heat, producing degradation and inactive side products, there is a need for a labeling technology performed under milder (e.g., room temperature) conditions, that avoids these conventional harsh labeling conditions, and can be completed rapidly in the hospital clinic prior to patient administration. Rapid labeling in a clinical setting is particularly important since many patients require diagnostic information quickly because of the acute nature of their condition.
Another class of chelating compounds developed for labeling targeting molecules are the bisamine bisthiols (termed BATs).
Baidoo et al., U.S. Pat. Nos. 5,196,515 and 5,095,111 disclose bisamine bisthiol complexes.
Kung et al., European Application No. 86105920.2 disclosed bisamine bisthiol ligands and their technetium-99m complexes.
Misra et al., 1989, Tet. Lett. 30: 1885–1888 describe bisamine bisthiol compounds for radiolabeling purposes.
Baidoo et al., 1990, Bioconjugate Chem. 1: 132–137 describe a method for labeling biomolecules using a bisamine bisthiol.
These compounds are useful when attached to targeting molecules since they can be labeled with technetium at room temperature. Such mild labeling conditions expose chemically-sensitive targeting molecules to a minimum of chemical stress, resulting in less degradation and more chemically pure radiolabeled targeting compounds. However, BAT chelates also have several drawbacks. One drawback of BAT chelators is that these chelates are intrinsically highly lipophilic. This property can cause these compounds to be retained in peripheral blood in excess, interfering with efficient scintigraphy because imaging agents must clear from the peripheral blood to reduce background radioactivity before a useful diagnostic image can be obtained. This drawback may alone be important enough to determine whether a BAT chelator-containing scintigraphic imaging agent is a commercially feasible product.
Another drawback of BAT chelators is that it is difficult to develop the chemistry to covalently attach such chelates to the targeting molecules. Although successful covalent linkage of BAT chelators to targeting molecules has been achieved, it has also typically resulted in the production of costly intermediates and has proven ultimately to be a costly way to produce the final radiopharmaceutical product.
The use of chelating agents for radiolabeling peptides, and methods for labeling peptides with Tc-99m are known in the prior art and are disclosed in co-pending U.S. patent application Ser. Nos. 07/653,012, 07/807,062, 07/871,282, 07/886,752, 07/893,981, 07/955,466, 08/019,864, 08/073,577, 08/210,822, 08/236,402 and 08/241,625, and radiolabeled peptides for use as scintigraphic imaging agents for imaging thrombi are known in the prior art and are disclosed in co-pending U.S. patent application Ser. Nos. 07/886,752, 07/893,981 and 08/044,825 and International Patent Applications Serial Nos. PCT/US92/00757, PCT/US92/10716, PCT/US93/02320, PCT/US93/03687, PCT/US93/04794, PCT/US93/05372, PCT/US93/06029, PCT/US93/09387, PCT/US94/01894, PCT/US94/03878, and PCT/US94/05895, each of which are hereby incorporated by reference in its entirety.
There exists a need for radiopharmaceuticals for diagnostic and therapeutic purposes that can be easily radiolabeled under mild chemical conditions to avoid chemical and physical degradation of labile biological targeting molecules. There remains a need for low-cost chelating groups which are easy to synthesize, moderately lipophilic, and can be linked to a targeting molecule and subsequently labeled with Tc-99m quickly at room temperature.